MTA2 Enhances Breast Cancer Metastasis by Regulating the Rho Signaling Pathway.

Abstract

Abstract Background: Metastasis is the ultimate cause of death for patients succumbing to breast cancer. The metastasis tumor associated (MTA) gene family has been linked with metastasis in breast cancer. Previously, we found that MTA2 overexpression enhanced the metastatic ability of the estrogen receptor-alpha (ERα)-negative cell line MDA-MB-231. This was accompanied by an increase in cell rounding, indicating a change in cytoskeletal organization. Here we explored the molecular pathways contributing to the MTA2-enhanced metastatic phenotype.Materials and Methods: MDA-MB-231 cells were engineered to overexpress MTA2. Changes in protein expression were determined by immunoblot assay. Differential gene expression was determined using Affymetrix microarrays and analyzed using dChip, BRB Array Tools, and R. Pathway analysis was carried out using the Database for Visualization, Annotation, and Integrated Discovery (DAVID). Validation was done by comparing to publicly available datasets. MTA2-overexpressing or control cells were transfected with a Rho GTP dissociation inhibitor alpha (Rho GDIα) or vector control expression constructs. Anchorage-independent growth was measured using soft-agar assays. Analysis of the impact of MTA2 and Rho GDIα combined expression was carried out using publically available datasets.Results: Analysis of enriched ontologies from significantly different genes between MTA2-overexpressing cells and clinical breast cancer samples of the basal subtype previously published by Richardson et al (1) indicated that the focal adhesion pathway was enhanced in MTA2-overexpressing samples. We found that a negative regulator of the focal adhesion pathway, Rho GDIα, was decreased in MTA2-overexpressing cells at the protein level. When a yellow fluorescent protein-tagged Rho GDIα expression vector was re-expressed in MTA2-overexpressing cells, there was a concomitant decrease in the expression of MTA2 and increased expression of endogenous Rho GDIα. When tested in soft agar assays, re-expression of Rho GDIα significantly reduced the colony-forming ability of MTA2-overexpressing cells. We examined the publically available dataset published by Wang et al. (2), and found that ERα-negative patients with high MTA2 expression and low Rho GDIa levels had earlier recurrence compared with all other patients (P=0.02).Conclusion: MTA2 overexpression was associated with an aggressive phenotype in MDA-MB-231 cells. These data indicate that MTA2 overexpression may be associated with increased signaling through the Rho pathway. We found that MTA2 and Rho GDIα form a regulatory loop which reinforces either MTA2 high or MTA2 low expression levels. Furthermore, we found that combined high MTA2 and low Rho GDIα expression had a negative impact on patient outcomes in the ERα-negative population. We are currently exploring the use of Rho pathway inhibitors for their potential to block this phenotype, and to explore the MTA2/Rho GDIα regulatory loop.1) Richardson AL, Wang ZC, De Nicolo A, Lu X et al. X chromosomal abnormalities in basal-like human breast cancer. Cancer Cell 2006 Feb;9(2):121-32.2) Wang Y, Klijn JG, Zhang Y, Sieuwerts AM et al. Gene-expression profiles to predict distant metastasis of lymph-node-negative primary breast cancer. Lancet 2005 Feb 19-25;365(9460):671-9. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 6143.